bims-unfpre 2025-11-23 papers

bims-unfpre

Biomed News

on Unfolded protein response

Issue of 2025–11–23
eight papers selected by
Susan Logue, University of Manitoba

CK2α Regulates Endoplasmic Reticulum Stress-Mediated Mitophagy via the PERK/ATF4/CHOP Pathway in Rotenone-Treated SH-SY5Y Cells.
From stress to homeostasis: Mass spectrometry-based insights into the unfolded protein response (UPR) and proteostasis.
The canonical ER stress IRE1α/XBP1 pathway mediates skeletal muscle wasting during pancreatic cancer cachexia.
Disrupted glycosylphosphatidylinositol anchoring induces ER stress and restricts enterovirus infection.
Design, Synthesis, and Cellular Efficacy of Inositol-Requiring Enzyme Type 1 (IRE1α) Inhibitors.
Inhibiting cholesterol synthesis halts rhabdomyosarcoma growth via ER stress and cell cycle arrest.
Inhibition of IRE1α by KIRA8 restores spingosine-1-phosphate lyase activity and improves insulin sensitivity in muscle cells.
RAF inhibitors activate the integrated stress response by direct activation of GCN2.

Mol Neurobiol. 2025 Nov 19. 63(1): 69

CK2α Regulates Endoplasmic Reticulum Stress-Mediated Mitophagy via the PERK/ATF4/CHOP Pathway in Rotenone-Treated SH-SY5Y Cells.

Kyung Mi Lim, Jungwook Hwang, Hyun Chul Koh.

  Mitophagy refers to selective mitochondrial autophagy to remove damaged mitochondria and plays a critical role in maintaining mitochondria homeostasis. Casein kinase 2α (CK2α) is involved in mitophagy regulation in dopaminergic neurons. Endoplasmic reticulum (ER) stress releases calcium into mitochondria, leading to mitochondrial dysfunction and contributing to various diseases. However, it is not clear whether CK2α regulates ER-mediated mitochondrial dysfunction and mitophagy in response to ER stress. Therefore, we investigated the effects of ER stress on mitophagy during rotenone-induced ER stress and mitochondrial damage in SH-SY5Y cells and elucidated the role of CK2α in this process. Rotenone increased the expression of P-PERK and P-IRE1α, thereby activating ER stress sensors. CK2α inhibition suppressed PERK and IRE1α activation and their downstream signaling components (eIF2α, ATF4, CHOP and XBP1s). Furthermore, CK2α inhibition enhanced PINK1/Parkin-mediated mitophagy by increasing PINK1 and Parkin translocation to mitochondria in addition to inducing LC3II expression. These results suggest that CK2α regulates PINK/Parkin-dependent mitophagy in rotenone-treated cells. Interestingly, treatment of cells with the PERK inhibitor GSK2606414 also resulted in increased PINK1/Parkin-mediated mitophagy. Moreover, CK2α inhibition reduced rotenone-induced apoptosis by modulating PERK signaling. These findings suggest that CK2α plays a key role in regulating the ER stress response and PERK-dependent PINK1/Parkin-mediated mitophagy in our rotenone-induced apoptosis model. This study highlights the therapeutic potential of CK2α signal regulation for treating diseases driven by ER stress and mitochondrial dysfunction, offering a promising avenue for future research.

Keywords:  Casein kinase 2α; ER stress; Mitophagy; PERK/ATF4/CHOP pathway; PINK/Parkin; Rotenone

DOI:  https://doi.org/10.1007/s12035-025-05441-z
Adv Biol Regul. 2025 Oct 30. pii: S2212-4926(25)00054-5. [Epub ahead of print] 101127

From stress to homeostasis: Mass spectrometry-based insights into the unfolded protein response (UPR) and proteostasis.

Lea A Barny, Lars Plate.

  The unfolded protein response (UPR) is a central regulator of proteostasis, coordinating cellular adaptation to endoplasmic reticulum (ER) stress. It is comprised of three signaling branches: ATF6 (activating transcription factor 6), IRE1 (inositol-requiring enzyme 1), and PERK (protein kinase RNA-like ER kinase), which mediate transcriptional and translational reprogramming of the proteostasis network. These pathways display both functional redundancy and branch-specific activities. Dysregulated UPR signaling contributes to diverse pathologies: in cancer, UPR activation supports uncontrolled proliferation and treatment resistance, whereas in aging, proteostasis decline and diminished UPR responsiveness are hallmarks. Traditional approaches, including transcriptomics and western blotting, have been widely used to monitor UPR activity, but they offer limited insight into its regulation at the protein level. In contrast, liquid chromatography-tandem mass spectrometry (LC-MS/MS) based proteomics allows comprehensive, branch-specific profiling of UPR signaling. Recent advances, including data-independent acquisition (DIA) MS and automated sample preparation, have further improved sensitivity, reproducibility, and detection of low-abundance UPR target proteins. Proteomics thus provides a systematic and scalable framework to interrogate UPR regulation across cell types and disease models. When integrated with complementary datasets, protein-level measurements can uncover context-dependent molecular signatures of UPR activity, offering insights into disease mechanisms and guiding the rational design of targeted pharmacological interventions. Future work integrating high-resolution LC-MS/MS proteomics with tissue and single-cell analyses will further clarify the role of the UPR in health and disease.

Keywords:  Activating transcription factor 6; Bottom-up proteomics; ER stress; Inositol requiring enzyme 1; Protein kinase R-like ER kinase; Proteomics automation; Proteostasis

DOI:  https://doi.org/10.1016/j.jbior.2025.101127
EMBO Mol Med. 2025 Nov 17.

The canonical ER stress IRE1α/XBP1 pathway mediates skeletal muscle wasting during pancreatic cancer cachexia.

Aniket S Joshi, Meiricris Tomaz da Silva, Anh Tuan Vuong, Bowen Xu, Ravi K Singh, Ashok Kumar.

  Cancer cachexia is a debilitating syndrome characterized by the progressive loss of skeletal muscle mass with or without fat loss. Recent studies have implicated dysregulation of the endoplasmic reticulum (ER) stress-induced unfolded protein response (UPR) pathways in skeletal muscle under various conditions, including cancer. In this study, we demonstrate that the IRE1α/XBP1 branch of the UPR promotes activation of the ubiquitin-proteasome system, autophagy, JAK-STAT3 signaling, and fatty acid metabolism in the skeletal muscle of the KPC mouse model of pancreatic cancer cachexia. Moreover, we show that the IRE1α/XBP1 pathway is a key contributor to muscle wasting. Skeletal muscle-specific deletion of the XBP1 transcription factor significantly attenuates tumor-induced muscle atrophy. Mechanistically, transcriptionally active XBP1 binds to the promoter regions of genes such as Map1lc3b, Fbxo32, and Il6, which encode proteins known to drive muscle proteolysis. Pharmacological inhibition of IRE1α using 4µ8C in KPC tumor-bearing mice attenuates cachexia-associated molecular changes and improves muscle mass and strength. Collectively, our findings suggest that targeting IRE1α/XBP1 pathway may offer a therapeutic strategy to counteract muscle wasting during pancreatic cancer-induced cachexia.

Keywords:  ER Stress; Fatty Acid Oxidation; JAK-STAT; Muscle Wasting; Unfolded Protein Response

DOI:  https://doi.org/10.1038/s44321-025-00337-w
PLoS Pathog. 2025 Nov;21(11): e1013685

Disrupted glycosylphosphatidylinositol anchoring induces ER stress and restricts enterovirus infection.

Shangrui Guo, Xinyu Li, Meng Xun, Yingli He, Andrew W Tai, Hongliang Wang.

Many positive-sense RNA viruses, including viruses from the Picornaviridae, Coronaviridae and Flaviviridae family, exploit endoplasmic reticulum (ER)-derived membrane structures as sites of genome replication. Here we use a pooled CRISPR genetic screening strategy to identify glycosylphosphatidylinositol (GPI) anchor biosynthesis and transfer genes as host factors for echovirus 7 infection. In addition to supporting the biogenesis of CD55, which is a GPI anchor protein and an entry factor for some echoviruses, the GPI anchor synthesis machinery also supports several other enterovirus infections by enhancing viral replication and replication organelle biogenesis. Disruption of GPI anchor transfer machinery compromises ER integrity and causes ER stress. Consistent with these findings, ER-resident sensor, inositol-requiring protein 1α (IRE1α) is activated and regulated IRE1-dependent decay of mRNA (RIDD) is detected to reduce ER stress. Interestingly, enterovirus viral RNA, but not Hepatitis C Virus RNA, is degraded during this process due to specific sequences in the Untranslated Region (UTR). This study revealed novel links between GPI anchoring, ER stress and enterovirus infection, and illuminates new host targets for antiviral therapy.

DOI: https://doi.org/10.1371/journal.ppat.1013685
ACS Med Chem Lett. 2025 Nov 13. 16(11): 2224-2231

Design, Synthesis, and Cellular Efficacy of Inositol-Requiring Enzyme Type 1 (IRE1α) Inhibitors.

Giulia Murbach, Adam D Mitrevski, Patricia A Fontan, Marcelo M Nociari, David H Thompson.

  A library of 66 small molecules targeting IRE1α were designed using a molecular docking approach and prepared by a two-step reaction sequence using diverse substrates. All compounds utilized a 1-amino-4-bromonaphthalene core that was modified via Suzuki coupling with boronic acids to form intermediates that were carbamoylated to form urea-linked inhibitor candidates. We developed a 33 DoE approach for the Suzuki coupling reaction that was optimized with 216 reactions via HTE. By screening the purified compounds in a tunicamycin-induced ER stress assay with ARPE-19 cells and quantifying their kinase inhibition activity by RT-qPCR, we identified 14 derivatives with the potential for IRE1α inhibition. IC50 assays showed that six of the compounds displayed IRE1α inhibition alike KIRA6, a standard in IRE1α inhibition, with three of the leads possessing improved IC50. Viability screens indicated that the best IRE1α inhibitors were not cytotoxic in the working concentrations and displayed improved protection from apoptosis compared to KIRA6.

Keywords:  ARPE-19; DESI-MS; DoE; ER stress; HTE; IRE1α inhibitors; KIRA6; Molecular docking; Suzuki coupling

DOI:  https://doi.org/10.1021/acsmedchemlett.5c00418
EMBO Mol Med. 2025 Nov 17.

Inhibiting cholesterol synthesis halts rhabdomyosarcoma growth via ER stress and cell cycle arrest.

Nebeyu Yosef Gizaw, Kalle Kolari, Pauliina Kallio, Kari Alitalo, Riikka Kivelä.

  Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma, with poor outcomes in high-risk and relapsed patients. Here, we identify de novo cholesterol biosynthesis as a critical metabolic vulnerability in RMS. The transcription factor PROX1, previously implicated in RMS growth, acts as an upstream regulator of cholesterol biosynthesis, promoting expression of key pathway genes. Inhibition of cholesterol biosynthesis, either genetically or pharmacologically, impaired RMS cell proliferation, caused a broad halt of cell cycle progression, and activated ER stress-mediated apoptosis through the PERK-ATF4-CHOP axis. Notably, RMS cells could not be rescued by exogenous LDL cholesterol, indicating a unique reliance on endogenous cholesterol production, whereas normal cells, including myoblasts and astrocytes, largely relied on extracellular cholesterol uptake. Clinical and single-cell RNA-seq analyses further revealed that high expression of cholesterol biosynthesis genes correlate with poor survival and enrichment of cell cycle-related gene signatures across RMS subtypes. Together, these findings mechanistically link cholesterol biosynthesis to proliferative signaling and ER stress response in RMS and highlight this pathway as a promising, non-redundant therapeutic target.

Keywords:  Drug Repurposing; Mevalonate Pathway; Pediatric Cancer; Sarcoma; Statins

DOI:  https://doi.org/10.1038/s44321-025-00336-x
Life Sci. 2025 Nov 16. pii: S0024-3205(25)00727-1. [Epub ahead of print]384 124091

Inhibition of IRE1α by KIRA8 restores spingosine-1-phosphate lyase activity and improves insulin sensitivity in muscle cells.

Javokhir Rustamov, Nodir Rustamov, Yuan-Qiang Ma, Yoon-Seok Roh, Tack-Joong Kim, Hwan-Soo Yoo.

   AIMS: Sphingosine-1-phosphate lyase (S1PL), a key enzyme in sphingolipid degradation, has been implicated in insulin resistance and type 2 diabetes. This study aimed to determine whether inhibition of IRE1α phosphorylation by KIRA8 during ER stress could restore S1PL activity, improve insulin signaling in C2C12 myotubes, and alleviate high-fat diet (HFD)-induced insulin resistance in mice.
MATERIALS AND METHODS: C2C12 myotubes were treated with palmitate (PA) or tunicamycin to induce ER stress. Effects of KIRA8 on pAkt, pIRE1α, glucose uptake, and S1PL activity were assessed. S1PL expression was modulated via Sgpl1 overexpression. Male C57BL/6N mice were fed a normal chow diet (NCD) or HFD for 10 weeks; KIRA8 was administered intraperitoneally to HFD-fed mice. Body weight, glucose and insulin tolerance, and histological changes in adipose tissue and liver were evaluated. pIRE1α, S1PL expression, and activity were measured in soleus muscle.
RESULTS: PA and tunicamycin reduced Akt phosphorylation and glucose uptake, increased pIRE1α, and inhibited S1PL activity all of which were reversed by KIRA8-mediated inhibition of pIRE1α in C2C12 myotubes. Sgpl1 overexpression enhanced S1PL activity and rescued insulin signaling. S1PL metabolites, phosphoethanolamine and hexadecenal, improved pAkt in PA-treated cells. In vivo, KIRA8 reduced body weight, adipose mass and liver size, and improved glucose metabolism. It also restored S1PL expression and activity and reduced pIRE1α in muscle.
CONCLUSIONS: IRE1α activation inhibits S1PL and impairs insulin signaling. KIRA8 restores S1PL function and insulin sensitivity during ER stress, identifying the IRE1α-S1PL axis as a potential therapeutic target for type 2 diabetes.

Keywords:  C2C12 myotubes; High-fat diet; IRE1α; Insulin signaling; KIRA8; S1PL

DOI:  https://doi.org/10.1016/j.lfs.2025.124091
Nat Commun. 2025 Nov 17. 16(1): 10033

RAF inhibitors activate the integrated stress response by direct activation of GCN2.

Rebecca Gilley, Andrew M Kidger, Graham Neill, Eve Morrison, Paul Severson, Dominic P Byrne, Niall S Kenneth, Gideon Bollag, Chao Zhang, Taiana Maia de Oliveira, Patrick A Eyers, Richard Bayliss, Glenn R Masson, Simon J Cook.

Paradoxical activation of wild type RAF by chemical RAF inhibitors (RAFi) is a well-understood 'on-target' biological and clinical response. In this study, we show that a range of RAFi drive ERK1/2-independent activation of the Unfolded Protein Response (UPR), including expression of ATF4 and CHOP, that requires the translation initiation factor eIF2α. RAFi-induced ATF4 and CHOP expression was not reversed by inhibition of PERK, a known upstream activator of the eIF2α-dependent Integrated Stress Response (ISR). Rather, RAFi exposure activated GCN2, an alternate eIF2α kinase, leading to eIF2α-dependent (and ERK1/2-independent) ATF4 and CHOP expression. The GCN2 kinase inhibitor A-92, GCN2 RNAi, GCN2 knock-out or ISRIB (an eIF2α antagonist) all reversed RAFi-induced expression of ATF4 and CHOP indicating that RAFi require GCN2 to activate the ISR. RAFi also activated full-length recombinant GCN2 in vitro and in cells, generating a characteristic 'bell-shaped' concentration-response curve, reminiscent of RAFi-driven paradoxical activation of WT RAF dimers. Activation of the ISR by RAFi was abolished by a GCN2 kinase dead mutation. A M802A GCN2 gatekeeper mutant was activated at lower RAFi concentrations, demonstrating that RAFi bind directly to the GCN2 kinase domain; this is supported by mechanistic structural models of RAFi interaction with GCN2. Since the ISR is a critical pathway for determining cell survival or death, our observations may be relevant to the clinical use of RAFi, where paradoxical GCN2 activation is a previously unappreciated off-target effect that may modulate tumour cell responses.

DOI: https://doi.org/10.1038/s41467-025-65376-w